Pediatric Surgery International

, Volume 27, Issue 6, pp 573–577 | Cite as

Prenatal administration of retinoic acid upregulates connective tissue growth factor in the nitrofen CDH model

  • Elke Maria Ruttenstock
  • Takashi Doi
  • Jens Dingemann
  • Prem Puri
Original Article



Recent studies have suggested that retinoids may be involved in the molecular mechanisms of pulmonary hypoplasia (PH) in congenital diaphragmatic hernia (CDH). Connective tissue growth factor (CTGF) plays a key role in foetal lung development and remodelling during later gestation. CTGF knockout mice exhibit PH with similar characteristics to the human and nitrofen-induced PH. Prenatal administration of retinoic acid (RA) has been shown to stimulate alveologenesis in nitrofen-induced PH. In vitro studies have revealed that RA can induce CTGF gene expression. We hypothesized that pulmonary gene expression of CTGF is downregulated during the later stages of lung development, and that prenatal administration of RA upregulates CTGF in the nitrofen CDH model.


Pregnant rats were exposed to either olive oil or nitrofen on day 9 (D9) of gestation. RA was given intraperitoneally on D18, D19 and D20. Foetuses were harvested on D21 and divided into control, CDH, control + RA and CDH + RA group. Pulmonary CTGF gene and protein expression levels were determined using RT-PCR and immunohistochemistry.


On D21, CTGF relative mRNA expression levels were significantly downregulated in CDH group compared to controls. After RA treatment, expression levels of CTGF were significantly upregulated in CDH + RA and control + RA compared to the CDH group. Immunohistochemical studies confirmed these results.


Downregulation of pulmonary CTGF gene and protein expression during later stages of lung development may interfere with normal alveologenesis in the nitrofen CDH model. Upregulation of CTGF pulmonary gene expression after prenatal RA treatment may promote lung growth by promoting alveologenesis in the nitrofen-induced CDH model.


Nitrofen Pulmonary hypoplasia (PH) Congenital diaphragmatic hernia (CDH) Retinoic acid (RA) Connective tissue growth factor (CTGF) 


  1. 1.
    Bohn D (2002) Congenital diaphragmatic hernia. Am J Respir Crit Care Med 166:911PubMedCrossRefGoogle Scholar
  2. 2.
    Stage G, Fenton A, Jaffray B (2003) Nihilism in the 1990s: the true mortality of congenital diaphragmatic hernia. Pediatrics 112:532–535CrossRefGoogle Scholar
  3. 3.
    Grosche JR, Islam S, Boulanger SC (2005) Congenital diaphragmatic hernia. Am J Surg 190:324–332CrossRefGoogle Scholar
  4. 4.
    Tenbrick R, Tibboel D, Gaillard J et al (1990) Experimentally induced congenital diaphragmatic hernia in rats. J Pediatr Surg 25:426–429CrossRefGoogle Scholar
  5. 5.
    Noble B, Babiuk R, Clugston R et al (2007) Mechanisms of congenital diaphragmatic hernia-inducing teratogen nitrofen. Am J Physiol Lung Cell Mol Physiol 293:1079–1087CrossRefGoogle Scholar
  6. 6.
    Antipatis C, Ashworth CJ, Grant G et al (1998) Effects of maternal vitamin A status on fetal heart and lung: changes in expression of key developmental genes. Am J Physiol Lung Cell Mol Physiol 275:1184–1191Google Scholar
  7. 7.
    Nakazawa N, Takayasu H, Montedonico S et al (2007) Altered regulation of retinoic acid synthesis in nitrofen-induced hypoplastic lung. Pediatr Surg Int 23:391–396PubMedCrossRefGoogle Scholar
  8. 8.
    Thebaud B, Tibboel D, Rambaud C et al (1999) Vitamin A decreases the incidence and severity of nitrofen-induced hypoplastic foetal rat lung explants. Pediatr Surg Int 277:423–429Google Scholar
  9. 9.
    Montedonico S, Nakazawa N, Puri P (2006) Retinoic acid rescues lung hypoplasia in the nitrofen-induced hypoplastic foetal rat lung explants. Pediatr Surg Int 22:2–8PubMedCrossRefGoogle Scholar
  10. 10.
    Montedonico S, Sugimoto K, Felle P et al (2008) Prenatal treatment with retinoic acid promotes pulmonary alveologenesis in the nitrofen model of congenital diaphragmatic hernia. J Pediatr Surg 43:500–507PubMedCrossRefGoogle Scholar
  11. 11.
    Sugimoto K, Takayasu H, Nakazawa N et al (2008) Prenatal treatment with retinoic acid accelerates type 1 alveolar cell proliferation of the hypoplastic lung in the nitrofen model of congenital diaphragmatic hernia. J Pediatr Surg 43:367–372PubMedCrossRefGoogle Scholar
  12. 12.
    Doi T, Sugimoto K, Puri P (2009) Prenatal retinoic acid up-regulates pulmonary gene expression of Coup-TFII, Fog2, and Gata4 in pulmonary hypoplasia. J Pediatr Surg 44:1933–1937PubMedCrossRefGoogle Scholar
  13. 13.
    Doi T, Sugimoto K, Ruttenstock E et al. (2010) Prenatal retinoic acid upregulates pulmonary gene expression of PI3K and AKT in nitrofen-induced pulmonary hypoplasia. Pediatr Surg Int (Epub ahead)Google Scholar
  14. 14.
    Surveyor G, Brigstock DR (1999) Immunohistochemical localization of connective tissue growth factor (CTGF) in the mouse embryo between days 7.5 and 14.5 of gestation. Growth Factors 17:115–124PubMedCrossRefGoogle Scholar
  15. 15.
    Blom IE, Goldschmeding R, Leask A (2002) Gene regulation of connective tissue growth factor: new targets for antifibrotic therapy. Matrix Biol 21:473–482PubMedCrossRefGoogle Scholar
  16. 16.
    Mesas-Burgos C, Nord M, Didon L et al (2009) Gene expression analysis after prenatal tracheal ligation in fetal rats as a model of stimulated lung growth. J Pediatr Surg 44:720–728PubMedCrossRefGoogle Scholar
  17. 17.
    Baguma-Nibasheka M, Kablar B (2008) Pulmonary hypoplasia in the connective tissue growth factor (Ctgf) null mouse. Dev Dyn 237:485–493PubMedCrossRefGoogle Scholar
  18. 18.
    Fadloun A, Kobi D, Delacroix L et al (2008) Retinoic acid induces TGFβ-dependent autocrine fribroblast growth. Oncogene 27:477–489PubMedCrossRefGoogle Scholar
  19. 19.
    Masumoto K, De Rooij J, Suita S et al (2006) The distribution of matrix metalloproteinases and tissue inhibitors of metalloproteinases in the lungs of congenital diaphragmatic hernia patients and age-matched controls. Histopathology 48:588–595PubMedCrossRefGoogle Scholar
  20. 20.
    Maden M, Hind M (2004) Retinoic acid in alveolar development, maintenance and regeneration. Philos Trans R Soc Lond B Biol Sci 359:799–808PubMedCrossRefGoogle Scholar
  21. 21.
    Massaro GD, Massaro D (2000) Retinoic acid treatment partially rescues failed septation in rats and mice. Am J Physiol 278:955–960Google Scholar
  22. 22.
    Nakazawa N, Montedonico S, Takayasu H et al (2007) Disturbance of retinol transportation causes nitrofen-induced hypoplastic lung. J Pediatr Surg 42:345–349PubMedCrossRefGoogle Scholar
  23. 23.
    Ruttenstock E, Doi T, Dingemann J et al (2010) Prenatal retinoic acid treatment upregulates late gestation lung protein 1 (LGL1) in the nitrofen-induced hypoplastic lung in late gestation. Pediatr Surg Int [Epub ahead of print]Google Scholar
  24. 24.
    Chang R, Andreoli S, Ng YS et al (2004) VEGF expression is downregulated in the nitrofen-induced congenital diaphragmatic hernia. J Pediatr Surg 39:825–828PubMedCrossRefGoogle Scholar
  25. 25.
    Oue T, Shima H, Taira Y et al (2000) Administration of antenatal glucocorticoids upregulates peptide growth factor gene expression in the nitrofen-induced congenital diaphragmatic hernia in rats. J Pediatr Surg 35:109–112PubMedCrossRefGoogle Scholar
  26. 26.
    Doi T, Ruttenstock E, Dingemann J et al (2010) Spatiotemporal alteration in phosphatidylinositide 3-kinase-serine/threonine protein kinase B signaling in the nitrofen-induced hypoplastic lung. J Pediatr Surg 45:366–371PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • Elke Maria Ruttenstock
    • 1
  • Takashi Doi
    • 1
    • 2
  • Jens Dingemann
    • 1
  • Prem Puri
    • 1
    • 2
  1. 1.National Children’s Research CentreOur Lady’s Children’s HospitalDublin 12Ireland
  2. 2.School of Medicine and Medical Science, Conway Institute of Biomolecular and Biomedical ResearchUniversity College DublinDublinIreland

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